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Proteomic Analysis Demonstrates Activator- and Chromatin-specific Recruitment to Promoters
Sikorski, Timothy W.,Joo, Yoo Jin,Ficarro, Scott B.,Askenazi, Manor,Buratowski, Stephen,Marto, Jarrod A. American Society for Biochemistry and Molecular Bi 2012 The Journal of biological chemistry Vol.287 No.42
<P>In-depth characterization of RNA polymerase II preinitiation complexes remains an important and challenging goal. We used quantitative mass spectrometry to explore context-dependent <I>Saccharomyces cerevisiae</I> preinitiation complex formation at the <I>HIS4</I> promoter reconstituted on naked and chromatinized DNA templates. The transcription activators Gal4-VP16 and Gal4-Gcn4 recruited a limited set of chromatin-related coactivator complexes, namely the chromatin remodeler Swi/Snf and histone acetyltransferases SAGA and NuA4, suggesting that transcription stimulation is mediated through these factors. Moreover, the two activators differentially recruited the coactivator complexes, consistent with specific activator-coactivator interactions. Chromatinized templates suppressed recruitment of basal transcription factors, thereby amplifying the effect of activators, compared with naked DNA templates. This system is sensitive, highly reproducible, and easily applicable to mapping the repertoire of proteins found at any promoter.</P>